Introduction
Assays | Technique | Scientific readout | Advantages | Disadvantages | |
---|---|---|---|---|---|
In vitro | Endothelial cell proliferation assay | Cell counting | Percentage of viable cells | Quantification of proliferating cells, apoptotic cells, and DNA content of the cell | Prone to manual error |
MTT assay | Measuring succinate dehydrogenase activity | Less expensive | Time-consuming | ||
3H-Thymidine /Bromouridine incorporation assay | DNA/RNA synthesis | ||||
Endothelial cell migration assay | Boyden chamber | Cell migration and invasion | Determination of migratory capacity of cells | Technical difficulties | |
Matrix degradation assay | Molecular factors influencing migration | Quantification of the rate of migration | Time-consuming | ||
Wound healing | Directional cell migration | Low rate of reproducibility | |||
Unable to accurately determine differences between proliferation and migration | |||||
Endothelial cell differentiation assay | Matrigel assay | Rearrangement of cells to form tubules | Quantification of pro-angiogenic factors | Technical difficulty | |
3D spheroid assays | Paracrine interactions and modulated pathways | Influence of biomolecules on ECs | Time-consuming | ||
Co-culturing ECs with other cell types | |||||
In vivo | Matrigel plug assay | Immuno-histochemistry staining | Quantification of newly formed blood vessels | Ideal model to study tissue regeneration | Expensive |
Time-consuming | |||||
CAM assay | Immuno-histochemistry staining | Formation of new blood vessels | Evaluation of angiogenic response | Sensitivity of the membrane to oxygen tension | |
Corneal angiogenesis assay | Microscopic observation | Vessel length and vascular sprouts | The reliable method as the cornea is devoid of pre-existing vasculature | Inappropriate for large scale studies | |
Immuno-histochemistry staining | |||||
Rodent mesentery angiogenesis assay | Immuno-histochemistry staining | Percentage of vascularized area | Extremely thin tissue enables easy visualization | Difficulty in quantification of angiogenesis | |
High sensitivity | |||||
Ex vivo | Rat aortic ring assay | Microscopic observation | Angiogenic sprouts and vessel length | Mimics in vivo conditions | Vessel growth is influenced by surrounding tissue |
Chick aortic arch assay | Microscopic observation | Cellular proliferation, migration, tube formation and vessel branching | Less expensive and less experimental time | Vessel growth is influenced by surrounding tissue | |
Rodent ear angiogenesis assay | Intravascular staining with biotinylated lectin | Vessel growth and branching | Easy visualization | Vessel growth is influenced by host cell interactions | |
Mimics in vivo conditions | |||||
Mouse fetal metatarsal angiogenesis assay | Immuno-histochemistry | Vessel sprouts, molecules influencing angiogenesis | Better representative of in vivo sprouts | Devoid of biomechanical force influencing the phenotype | |
Employs microvascular cells | Requires technical precision |
Mechanical stimulation | ||||
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Biomaterial | Cell type | Strategy | Application | Reference |
Collagen hydrogel | Rat microvessel fragments | Static external loader cyclic external load | Angiogenic microvessel | Krishnan et al. (2008) |
Fibrin gel | Human blood outgrowth endothelial cells | Cell-induced gel compaction | Aligned microvessels | |
Electrical stimulation | ||||
Biomaterial | Cell type | Strategy | Application | Reference |
Matrigel | Human mammary epithelial cells, HUVECs | DC electric field | Directional migration of cells | Bai et al. (2004) |
Surface topography | ||||
Biomaterial | Cell type | Strategy | Application | Reference |
Silk fibroin and fibrin | HUVECs + human foreskin fibroblasts | 3D porous scaffolds | Capillary‐like structure formation | Samal et al. (2015) |
Silk fibroin fibers in poly (d,l-lactic acid) porous scaffolds | Human endothelial cells | 3D salt-leached scaffolds | In vitro endothelial and to promote vascularization in vivo | Stoppato et al. (2013) |
Gelatin methacrylate hydrogels | Human blood-derived endothelial colony-forming cells and bone marrow-derived mesenchymal stem cells | 3D porous scaffolds | The functional human vascular network | Chen et al. (2012) |
Silk fibroin | Human microvascular endothelial and osteoblast cells | 3D fibrous scaffolds | Anastomosis of neo-microcapillaries with the host vasculature | Unger et al. 92010) |
Collagen | Endothelial colony-forming cells and endothelial progenitor cells | 3D fibrous scaffolds (varied collagen concentration) | Guiding in vivo vascularization | Critser et al. (2010) |
Decellularized fibroblasts derived ECM | Human mesenchymal stem cells | 3D nanofibrous scaffolds | Engineering organized tissues | Xing et al. (2014) |